Vegetative death or senescence occurs in all races of Podospora anserina. It has been known for quite some time that senescence is cytoplasmically inherited in intra and interracial crosses and that mitochondrial function is involved. Previously, we showed that mitochondrial (mt) DNA from young Podospora consists of 30 micron convalently closed circles. During senescence, at a time when the mitochondrial respiratory pathway is impaired, we made two observations: 1) A significant fraction (about 10%) of the mt DNA consists of a multimeric series of circles; no full length (30 micron) were observed although restriction enzyme analysis showed the presence of most DNA fragments but at different concentrations, and 2) in independent senescent events, the tandemly repeated monomeric units contained in the multimeric circles were of different sizes, ranging from 800 bp to 9000 bp. Many of these DNA fragments have been cloned. We have two major objectives in the coming year. A) To determine the relationship between young mt DNA and the mt DNA fragments issued from mt isolated from independent senescent cultures. This will include homology studies of the intact inserts from clones as well as the use of fine cutting restriction endonucleases. B) To ascertain whether any or all of of these DNA fragments will "transform" protoplasts from young cultures of Podospora to premature senescence. Should senescence result, we will then inquire whether the multimeric set of circles resulting will have the same monomeric repeat as the donor cloned insert or whether a different molecular event has been induced.